Diltiazem inhibits SARS-CoV-2 cell attachment and internalization and decreases the viral infection in mouse lung.

The continuous emergence of severe acute respiratory coronavirus 2 (SARS-CoV-2) variants and the increasing number of breakthrough infection cases among vaccinated people support the urgent need for research and development of antiviral drugs. Viral entry is an intriguing target for antiviral drug development. We found that diltiazem, a blocker of the L-type calcium channel Cav1.2 pore-forming subunit (Cav1.2 α1c) and an FDA-approved drug, inhibits the binding and internalization of SARS-CoV-2, and decreases SARS-CoV-2 infection in cells and mouse lung. Cav1.2 α1c interacts with SARS-CoV-2 spike protein and ACE2, and affects the attachment and internalization of SARS-CoV-2. Our finding suggests that diltiazem has potential as a drug against SARS-CoV-2 infection and that Cav1.2 α1c is a promising target for antiviral drug development for COVID-19.

Platelet-activating factor contracts guinea pig esophageal muscularis mucosae by stimulating extracellular Ca2+ influx through diltiazem-insensitive Ca2+ channels.

Platelet-activating factor (PAF) is expected to increase esophageal motility. However, to the best of our knowledge, this has not been examined. Thus, we investigated the contractile effects of PAF on guinea pig (GP) esophageal muscularis mucosae (EMM) and the extracellular Ca2+ influx pathways responsible. PAF (10-9-10-6 M) contracted EMM in a concentration-dependent manner. PAF (10-6 M)-induced contractions were almost completely suppressed by apafant (a PAF receptor antagonist, 3 × 10-5 M). In EMM strips, PAF receptor and PAF-synthesizing/degrading enzyme mRNAs were detected. PAF (10-6 M)-induced contractions were abolished by extracellular Ca2+ removal but were not affected by diltiazem [a voltage-dependent Ca2+ channel (VDCC) inhibitor, 10-5 M]. PAF (10-6 M)-induced contractions in the presence of diltiazem were significantly suppressed by LOE-908 [a receptor-operated Ca2+ channel (ROCC) inhibitor, 3 × 10-5 M], SKF-96365 [an ROCC and store-operated Ca2+ channel (SOCC) inhibitor, 3 × 10-5 M], and LOE-908 plus SKF-96365. Among the tested ROCC/SOCC-related mRNAs, Trpc3, Trpc6, and Trpv4/Orai1, Orai3, and Stim2 were abundantly expressed in EMM strips. These results indicate that PAF potently induces GP EMM contractions that are dependent on extracellular Ca2+ influx through ROCCs/SOCCs, and VDCCs are unlikely to be involved.

Relaxant Effects of Piperine and Piperlongumine from Piper longum Fruits on Porcine Coronary Artery.

Since ancient times, Piper longum Linn. fruits have been recognized for exhibiting various effects, including the diaphoretic effects linked to enhanced blood flow. Piperine and piperlongumine coexist in Piper longum Linn. fruits, although the cardiovascular effects of both compounds remain elusive. We investigated their action of piperine and piperlongumine in porcine coronary arteries, comparing them to the Ca2+ channel antagonist diltiazem. Piperlongumine, unlike piperine or diltiazem, concentration-dependently inhibited basal contractile tone in endothelium-denuded coronary arteries. All three compounds inhibit tonic contractions induced by high potassium chloride (KCl) concentrations. The order of relaxation potency indexed by the half-maximal effective concentration (EC50) were as follows: diltiazem > piperlongumine > piperine. These effects were not different between endothelium-intact and -denuded preparations. In endothelial-denuded preparations, pretreatment with these compounds not only inhibited KCl-induced tonic contractions attenuated calcium chloride (CaCl2)-induced ones in a Ca2+-free medium. Histamine-induced phasic contractions in a Ca2+-free medium containing intracellular Ca2+ chelator was completely suppressed by selective inositol trisphosphate receptor antagonist and piperlongumine, whereas piperine or diltiazem do not have the same effect. These findings suggest that piperine and piperlongumine similar to diltiazem cause vasorelaxation by inhibiting both KCl- and CaCl2-induced contractions in coronary arteries, possibly through the inhibition of voltage-dependent Ca2+ channels. Piperlongumine inhibits histamine-induced contractions in a Ca2+-free medium, which is associated with the intracellular Ca2+ signaling pathway, suggesting that the relaxant effect of piperlongumine differs from that of piperine.

Glucocorticoid effects on working memory impairment require l-type calcium channel activity within prefrontal cortex.

Previous findings have indicated that glucocorticoid hormones impair working memory via an interaction with the ß-adrenoceptor-cAMP signaling cascade to rapidly increase cAMP-dependent protein kinase (PKA) activity within the prefrontal cortex (PFC). However, it remains elusive how such activation of PKA can affect downstream cellular mechanisms in regulating PFC cognitive function. PKA is known to activate l-type voltage-gated Ca2+ channels (LTCCs) which regulate a broad range of cellular processes, including neuronal excitability and neurotransmitter release. The present experiments examined whether LTCC activity within the PFC is required in mediating glucocorticoid and PKA effects on spatial working memory. Male Sprague Dawley rats received bilateral administration of the LTCC inhibitor diltiazem together with either the glucocorticoid receptor agonist RU 28362 or PKA activator Sp-cAMPS into the PFC before testing on a delayed alternation task in a T-maze. Both RU 28362 and Sp-cAMPS impaired working memory, whereas the LTCC inhibitor diltiazem fully blocked the working memory impairment induced by either RU 28362 or Sp-cAMPS. Conversely, bilateral administration of the LTCC agonist Bay K8644 into the PFC was sufficient to impair working memory. Thus, these findings provide support for the view that glucocorticoids, via an interaction with the ß-adrenergic signaling cascade and enhanced PKA activity levels, impair working memory by increasing LTCC activity in the PFC.

In Esophageal Squamous Cells From Eosinophilic Esophagitis Patients, Th2 Cytokines Increase Eotaxin-3 Secretion Through Effects on Intracellular Calcium and a Non-Gastric Proton Pump.

BACKGROUND & AIMS: In upper airway cells, T helper 2 cytokines that signal through interleukin-4 (IL-4) receptor-α have been shown to stimulate eotaxin-3 secretion via a nongastric proton pump (ngH+,K+ATPase). To seek novel targets for eosinophilic esophagitis (EoE) treatments, we evaluated ngH+,K+ATPase expression in EoE squamous cells, and explored molecular pathways involved in eotaxin-3 secretion by IL-4 receptor-α signaling. METHODS: ngH+,K+ATPase expression in EoE cells was evaluated by quantitative real-time polymerase chain reaction and Western blotting. IL-4-stimulated eotaxin-3 secretion was measured by enzyme-linked immunosorbent assay after treatment with omeprazole, SCH 28080 (potassium-competitive acid blocker), ethylene glycol-bis(ß-aminoethyl)-N,N,N',N'-tetraacetoxymethyl ester (calcium chelator), 2-aminoethoxydiphenyl borate (inhibitor of endoplasmic reticulum calcium release), verapamil, and diltiazem (L-type calcium channel inhibitors). Intracellular calcium transients were measured by Fluo-4 fluorescence. Key experiments were confirmed in EoE primary cells and in RNA sequencing datasets from mucosal biopsies of patients with EoE and controls. RESULTS: EoE cells expressed ngH+,K+ATPase messenger RNA and protein. Omeprazole and SCH 28080 decreased IL-4-stimulated eotaxin-3 secretion. IL-4 increased intracellular calcium transients, and IL-4-stimulated eotaxin-3 secretion was blocked by ethylene glycol-bis(ß-aminoethyl)-N,N,N',N'-tetraacetoxymethyl ester, 2-aminoethoxydiphenyl borate, verapamil, and diltiazem. The combination of omeprazole and verapamil suppressed IL-4-stimulated eotaxin-3 secretion more than either agent alone. EoE biopsies expressed higher ngH+,K+ATPase and exhibited more calcium signaling than controls. CONCLUSIONS: EoE cells express a nongastric proton pump that mediates T helper 2 cytokine-stimulated eotaxin-3 secretion. IL-4 induces calcium release from the endoplasmic reticulum and calcium entry via L-type calcium channels, increasing intracellular calcium that contributes to eotaxin-3 secretion by EoE cells. L-type calcium channel inhibitors block T helper 2 cytokine-stimulated eotaxin-3 secretion, suggesting a potential role for these agents in EoE treatment.

In vitro and in vivo impacts of nifedipine and diltiazem on praziquantel chemotherapy in murine Schistosoma mansoni.

AIM: This study was planned to evaluate the in vitro and in vivo antischistosomal effects of the widely used antihypertensive drugs, nifedipine (NIF) and diltiazem (DTZ), and their combinations with praziquantel (PZQ) on early and late Schistosoma (S.) mansoni infections 21- and 45- days old stages. METHODS: In the In vitro study, Calcium channel blockers (CCBs), NIF and DTZ were added to schistosomula and adult worm cultures in different concentrations 10, 20 and 30 mg/ml. The mortality percentage was calculated 1, 12 and 24 h after incubation. In vivo, NIF and DTZ either alone or combined with PZQ were used to treat male albino mice. The parasitological and total immunoglobulin (Ig) G and IgM anti-soluble egg antigen (SEA) were assessed to demonstrate the disease severity. RESULTS: In the In vitro study, 10 mg/ml NIF induced 100% mortality percentage of both schistosomula and adult worms after 24 h incubation, while DTZ induced similar mortality percentage at 30 mg/ml concentration. In vivo results showed that early or late combination of 30 mg/kg of NIF, but not DTZ, significantly (P <0.05) enhanced the reductive efficacy of PZQ based on the parasitological data. The maximal reduction (P <0.05) of anti-SEA IgM and IgG levels was developed during NIF-PZQ administration 21- (1.12 ± 0.06 and 1.09 ± 0.04, respectively) or 45- (1.00 ± 0.03 and 0.8 ± 0.06, respectively) days post infection (PI), compared to either PZQ or NIF individual treatments. The decreased concentration of anti-SEA antibodies was correlated with the diminished granulomatous diameter and disease severity. CONCLUSION: Nifedipine improved PZQ chemotherapy targeting either early or late S. mansoni infection in mice compared to the PZQ mono-therapy. Administering NIF can be considered as a promising drug candidate for schistosomiasis chemotherapy.

[Effect and mechanism of Huangqi Shengmai Decoction in treatment of joint rat model of fatigue and myocardial injury].

This study aims to investigate the effects and the underlying mechanism of Huangqi Shengmai Decoction(HQSMD) in the treatment of fatigue and myocardial injury in a joint rat model. Wistar rats were assigned into 4 groups: sham, model, diltiazem hydrochloride(positive control), and HQSMD. The joint model of fatigue and myocardial injury was established by 14-day exhausted swimming followed by high ligation of the left anterior descending coronary artery. The rats in the sham group underwent a sham operation without coronary artery ligation or swimming. Since the fourth day after the ligation, swimming was continued in the model group and the drug-treated groups for the following 4 weeks. Meanwhile, the rats in the positive control group and the HQSMD group were respectively administrated intragastrically with diltiazem hydrochloride(20 mg·kg~(-1)·d~(-1)) and HQSMD(0.95 g·kg~(-1)·d~(-1)) for 4 weeks, while the shams and the models were given the same volume of normal saline. The left ventricular ejection fraction(LVEF), left ventricular fractional shortening(LVFS), grip strength, and myocardial pathophysiological changes were measured to evaluate the anti-fatigue and cardioprotective effects of HQSMD. The protein levels of PTEN-induced putative kinase 1(PINK1) and parkin in the myocardium were measured by Western blot to preliminarily elucidate the mechanism of HQSMD in ameliorating myocardial injury by suppressing mitochondrial autophagy. Compared with the shams, the models showed weakened heart function(LVEF and LVFS, P<0.01), decreased grasping ability(P<0.05), elevated blood urea nitrogen(BUN) and aldosterone(ALD) levels(P<0.01), aggravated myocardial fibrosis and connective tissue hyperplasia(P<0.01), and up-regulated protein levels of PINK1(P<0.01) and parkin(P<0.05). Four-week treatment with HQSMD increased the LVEF and LVFS levels(P<0.01), enhanced the grip strength(P<0.01), reduced the serum levels of BUN(P<0.01) and ALD(P<0.05), alleviated the pathological injury and fibrosis in the myocardium(P<0.01), and down-regulated the protein levels of PINK1(P<0.01) and parkin(P<0.05) in heart tissue. The results demonstrate that HQSMD may alleviate myocardial fibrosis and protect myocardium by suppressing the excessive mitochondrial auto-phagic activity and reducing the excessively elevated ALD level, thereby ameliorating fatigue and myocardial injury.

Construction and Characterization of CRISPR/Cas9 Knockout Rat Model of Carboxylesterase 2a Gene.

Carboxylesterase (CES) 2, an important metabolic enzyme, plays a critical role in drug biotransformation and lipid metabolism. Although CES2 is very important, few animal models have been generated to study its properties and functions. Rat Ces2 is similar to human CES2A-CES3A-CES4A gene cluster, with highly similar gene structure, function, and substrate. In this report, CRISPR-associated protein-9 (CRISPR/Cas9) technology was first used to knock out rat Ces2a, which is a main subtype of Ces2 mostly distributed in the liver and intestine. This model showed the absence of CES2A protein expression in the liver. Further pharmacokinetic studies of diltiazem, a typical substrate of CES2A, confirmed the loss of function of CES2A both in vivo and in vitro. At the same time, the expression of CES2C and CES2J protein in the liver decreased significantly. The body and liver weight of Ces2a knockout rats also increased, but the food intake did not change. Moreover, the deficiency of Ces2a led to obesity, insulin resistance, and liver fat accumulation, which are consistent with the symptoms of nonalcoholic fatty liver disease (NAFLD). Therefore, this rat model is not only a powerful tool to study drug metabolism mediated by CES2 but also a good disease model to study NAFLD. SIGNIFICANCE STATEMENT: Human carboxylesterase (CES) 2 plays a key role in the first-pass hydrolysis metabolism of most oral prodrugs as well as lipid metabolism. In this study, CRISPR/Cas9 technology was used to knock out Ces2a gene in rats for the first time. This model can be used not only in the study of drug metabolism and pharmacokinetics but also as a disease model of nonalcoholic fatty liver disease (NAFLD) and other metabolic disorders.

Identifying the Transcriptome Signatures of Calcium Channel Blockers in Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes.

RATIONALE: Calcium channel blockers (CCBs) are an important class of drugs in managing cardiovascular diseases. Patients usually rely on these medications for the remainder of their lives after diagnosis. Although the acute pharmacological actions of CCBs in the hearts are well-defined, little is known about the drug-specific effects on human cardiomyocyte transcriptomes and physiological alterations after long-term exposure. OBJECTIVE: This study aimed to simulate chronic CCB treatment and to examine both the functional and transcriptomic changes in human cardiomyocytes. METHODS AND RESULTS: We differentiated cardiomyocytes and generated engineered heart tissues from 3 human induced pluripotent stem cell lines and exposed them to 4 different CCBs-nifedipine, amlodipine, diltiazem, and verapamil-at their physiological serum concentrations for 2 weeks. Without inducing cell death and damage to myofilament structure, CCBs elicited line-specific inhibition on calcium kinetics and contractility. While all 4 CCBs exerted similar inhibition on calcium kinetics, verapamil applied the strongest inhibition on cardiomyocyte contractile function. By profiling cardiomyocyte transcriptome after CCB treatment, we identified little overlap in their transcriptome signatures. Verapamil is the only inhibitor that reduced the expression of contraction-related genes, such as MYH (myosin heavy chain) and troponin I, consistent with its depressive effects on contractile function. The reduction of these contraction-related genes may also explain the responsiveness of patients with hypertrophic cardiomyopathy to verapamil in managing left ventricular outflow tract obstruction. CONCLUSIONS: This is the first study to identify the transcriptome signatures of different CCBs in human cardiomyocytes. The distinct gene expression patterns suggest that although the 4 inhibitors act on the same target, they may have distinct effects on normal cardiac cell physiology.

Evaluation of metoprolol versus diltiazem for rate control of atrial fibrillation in the emergency department.

OBJECTIVE: The purpose of this study was to compare the effectiveness and safety of the metoprolol and diltiazem administration in the Emergency Department (ED) for rate control of supraventricular tachycardia. METHODS: This was a retrospective cohort study of adult patients who presented to the ED with ventricular rates ≥120 beats per minute (bpm) and who received bolus doses of either intravenous metoprolol or intravenous diltiazem. The primary outcome was achievement of rate control, defined as heart rate < 110 bpm, at two hours after administration of the last bolus dose of metoprolol or diltiazem. Safety outcomes included occurrence of hypotension, defined as systolic blood pressure < 90 mmHg or diastolic blood pressure < 60 mmHg, and bradycardia, defined as heart rate < 60 bpm. RESULTS: There were 166 patients receiving metoprolol and 183 patients receiving diltiazem included in the study. The primary outcome, rate control at two hours after the last bolus dose of metoprolol or diltiazem was similar between the two groups (45.8% vs 42.6%, p = 0.590, respectively). The percentage of patients achieving rate control was also similar (47.0% vs 41.6%, p = 0.333) at one hour. At 0.5 h HR had a significantly greater numerical (diltiazem: 29.3 ± 23.1 bpm vs metoprolol: 21.8 ± 18.9 bpm, p = 0.012) and percent decrease (21.1% vs 15.94%, p = 0.014) in the diltiazem group compared to metoprolol. There was no significant difference in occurrence of bradycardia in the two groups (diltiazem: 3.83% vs metoprolol: 1.2%, p = 0.179). More patients in the diltiazem group compared to the metoprolol group experienced hypotension (39.3% vs 23.5%, p = 0.002). The difference in systolic hypotension events was not significantly different (9.29% vs 5.42%, p = 0.221), while the difference in diastolic hypotension events was significantly different (37.7% vs 22.3%, p = 0.002). CONCLUSION: There was no difference in acute rate control effectiveness two hours after the last bolus dose of diltiazem and metoprolol for supraventricular tachycardias. There was a significantly higher occurrence of hypotension in the diltiazem group which was driven by higher rates of diastolic blood pressures less than 60 mmHg.

Role of L-Type Voltage-Gated Calcium Channels in Epileptiform Activity of Neurons.

Epileptic discharges manifest in individual neurons as abnormal membrane potential fluctuations called paroxysmal depolarization shift (PDS). PDSs can combine into clusters that are accompanied by synchronous oscillations of the intracellular Ca2+ concentration ([Ca2+]i) in neurons. Here, we investigate the contribution of L-type voltage-gated calcium channels (VGCC) to epileptiform activity induced in cultured hippocampal neurons by GABA(A)R antagonist, bicuculline. Using KCl-induced depolarization, we determined the optimal effective doses of the blockers. Dihydropyridines (nifedipine and isradipine) at concentrations ≤ 10 µM demonstrate greater selectivity than the blockers from other groups (phenylalkylamines and benzothiazepines). However, high doses of dihydropyridines evoke an irreversible increase in [Ca2+]i in neurons and astrocytes. In turn, verapamil and diltiazem selectively block L-type VGCC in the range of 1-10 µM, whereas high doses of these drugs block other types of VGCC. We show that L-type VGCC blockade decreases the half-width and amplitude of bicuculline-induced [Ca2+]i oscillations. We also observe a decrease in the number of PDSs in a cluster and cluster duration. However, the pattern of individual PDSs and the frequency of the cluster occurrence change insignificantly. Thus, our results demonstrate that L-type VGCC contributes to maintaining the required [Ca2+]i level during oscillations, which appears to determine the number of PDSs in the cluster.

Diltiazem Inhibits Coronary Spasm via Inhibition of Cav1.2Phosphorylation and Protein Kinase C Activation in a Mouse Model of Coronary Spastic Angina.

Calcium antagonists are used for coronary spastic angina (CSA) treatment. We previously identified a phospholipase C (PLC) -δ1 gene variant that results in enhanced PLC activity in patients with CSA and developed a CSA animal model by generating vascular smooth muscle cell-specific human variant PLC-δ1 overexpression (PLC-TG) mice. In this study, we investigated the molecular mechanism of CSA using the PLC-TG mice and the inhibitory effect of a calcium antagonist, diltiazem hydrochloride (DL).We treated the PLC-TG and wild-type (WT) mice with oral DL or trichlormethiazide (TM) (control) for 2 weeks. Ergometrine injection-induced coronary spasm was observed on the electrocardiogram in all 5 PLC-TG mice treated with TM, but only in 1 of 5 PLC-TG mice treated with DL. Voltage-dependent calcium channel (Cav1.2) phosphorylation and protein kinase C (PKC) activity were enhanced in the aortas of PLC-TG mice treated with TM. DL treatment significantly inhibited Cav1.2 phosphorylation and PKC activity. Although total Cav1.2 expression was similar between WT and PLC-TG mice treated with TM, DL treatment significantly increased its expression in PLC-TG mice. Furthermore, its expression remained high after DL discontinuation. DL and PKC inhibitor suppressed intracellular calcium response to acetylcholine in cultured rat aortic smooth muscle cells transfected with variant PLC-δ1.These results indicate that enhanced PLC activity causes coronary spasm, presumably via enhanced Cav1.2 phosphorylation and PKC activity, both of which were inhibited by DL. Enhanced total Cav1.2 expression after DL discontinuation and high PKC activity may be an important mechanism underlying the calcium antagonist withdrawal syndrome.

Diltiazem-loaded electrospun nanofibers as a new wound dressing: fabrication, characterization, and experimental wound healing.

Calcium channel blockers such as diltiazem have recently been investigated for their wound-healing potential. The aims of this study were to fabricate diltiazem-loaded nanofibers for a new wound dressing and investigate their beneficial properties for wound healing. Nanofibers were electrospun using polyvinyl alcohol solution containing 0, 2 or 4% diltiazem. Fibers were characterized in terms of physicochemical properties, drug release and fibroblast viability, and in animal wound healing assays. Compared to other formulations, nanofibers containing 4% diltiazem showed thin fiber size (152.7 nm), high porosity (88.4%), high swelling (110.4%), low water contact angle (29.1°) and little weight loss (17.3%). Drug release from 4%-diltiazem nanofibers showed good fit to a Korsmeyer-Peppas model, suggesting a non-Fickian release mechanism (R 2 = 96%, n = 0.52). In vitro, 4%-diltiazem mats were not cytotoxic and enhanced fibroblast proliferation by 263% after 5 days of treatment compared to control. In vivo, wounds treated with this mat for 14 days showed the smallest size (14.7%) and better histopathologic characteristics compared to other wounds. The 4%-diltiazem mat also demonstrated significant antioxidant activity by reducing tissue MDA and nitrite levels by 63 and 59% compared to normal saline. The findings support the eligibility of this novel wound dressing for additional clinical research.

Chronic Calmodulin-Kinase II Activation Drives Disease Progression in Mutation-Specific Hypertrophic Cardiomyopathy.

BACKGROUND: Although the genetic causes of hypertrophic cardiomyopathy (HCM) are widely recognized, considerable lag in the development of targeted therapeutics has limited interventions to symptom palliation. This is in part attributable to an incomplete understanding of how point mutations trigger pathogenic remodeling. As a further complication, similar mutations within sarcomeric genes can result in differential disease severity, highlighting the need to understand the mechanism of progression at the molecular level. One pathway commonly linked to HCM progression is calcium homeostasis dysregulation, though how specific mutations disrupt calcium homeostasis remains unclear. METHODS: To evaluate the effects of early intervention in calcium homeostasis, we used 2 mouse models of sarcomeric HCM (cardiac troponin T R92L and R92W) with differential myocellular calcium dysregulation and disease presentation. Two modes of intervention were tested: inhibition of the autoactivated calcium-dependent kinase (calmodulin kinase II [CaMKII]) via the AC3I peptide and diltiazem, an L-type calcium channel antagonist. Two-dimensional echocardiography was used to determine cardiac function and left ventricular remodeling, and atrial remodeling was monitored via atrial mass. Sarcoplasmic reticulum Ca2+ATPase activity was measured as an index of myocellular calcium handling and coupled to its regulation via the phosphorylation status of phospholamban. RESULTS: We measured an increase in phosphorylation of CaMKII in R92W animals by 6 months of age, indicating increased autonomous activity of the kinase in these animals. Inhibition of CaMKII led to recovery of diastolic function and partially blunted atrial remodeling in R92W mice. This improved function was coupled to increased sarcoplasmic reticulum Ca2+ATPase activity in the R92W animals despite reduction of CaMKII activation, likely indicating improvement in myocellular calcium handling. In contrast, inhibition of CaMKII in R92L animals led to worsened myocellular calcium handling, remodeling, and function. Diltiazem-HCl arrested diastolic dysfunction progression in R92W animals only, with no improvement in cardiac remodeling in either genotype. CONCLUSIONS: We propose a highly specific, mutation-dependent role of activated CaMKII in HCM progression and a precise therapeutic target for clinical management of HCM in selected cohorts. Moreover, the mutation-specific response elicited with diltiazem highlights the necessity to understand mutation-dependent progression at a molecular level to precisely intervene in disease progression.

Optical control of L-type Ca2+ channels using a diltiazem photoswitch.

L-type Ca2+ channels (LTCCs) play a crucial role in excitation-contraction coupling and release of hormones from secretory cells. They are targets of antihypertensive and antiarrhythmic drugs such as diltiazem. Here, we present a photoswitchable diltiazem, FHU-779, which can be used to reversibly block endogenous LTCCs by light. FHU-779 is as potent as diltiazem and can be used to place pancreatic ß-cell function and cardiac activity under optical control.

Benzothiazepines, diltiazem and JTV-519, exert an anxiolytic-like effect via neurosteroid biosynthesis in mice.

We investigated whether benzothiazepines could produce anxiolytic effects via allopregnanolone (ALLO) biosynthesis in mice. We compared the behavioral effects caused by benzothiazepines to those caused by carbamazepine and sodium valproate. We found that a pretreatment with finasteride (a 5 alpha-reductase inhibitor) suppressed carbamazepine-induced anxiolytic effects but not the effects of sodium valproate. Similar to carbamazepine, diltiazem and JTV-519 displayed anxiolytic effects that were suppressed by a pretreatment with finasteride. We clearly demonstrate that the benzothiazepines, diltiazem and JTV-519, exert an anxiolytic-like effect via ALLO biosynthesis in mice.

Impact of Electronic Health Record Interface Design on Unsafe Prescribing of Ciclosporin, Tacrolimus, and Diltiazem: Cohort Study in English National Health Service Primary Care.

BACKGROUND: In England, national safety guidance recommends that ciclosporin, tacrolimus, and diltiazem are prescribed by brand name due to their narrow therapeutic windows and, in the case of tacrolimus, to reduce the chance of organ transplantation rejection. Various small studies have shown that changes to electronic health record (EHR) system interfaces can affect prescribing choices. OBJECTIVE: Our objectives were to assess variation by EHR systems in breach of safety guidance around prescribing of ciclosporin, tacrolimus, and diltiazem, and to conduct user-interface research into the causes of such breaches. METHODS: We carried out a retrospective cohort study using prescribing data in English primary care. Participants were English general practices and their respective EHR systems. The main outcome measures were (1) the variation in ratio of safety breaches to adherent prescribing in all practices and (2) the description of observations of EHR system usage. RESULTS: A total of 2,575,411 prescriptions were issued in 2018 for ciclosporin, tacrolimus, and diltiazem (over 60 mg); of these, 316,119 prescriptions breached NHS guidance (12.27%). Breaches were most common among users of the EMIS EHR system (breaches in 18.81% of ciclosporin and tacrolimus prescriptions and in 17.99% of diltiazem prescriptions), but breaches were observed in all EHR systems. CONCLUSIONS: Design choices in EHR systems strongly influence safe prescribing of ciclosporin, tacrolimus, and diltiazem, and breaches are prevalent in general practices in England. We recommend that all EHR vendors review their systems to increase safe prescribing of these medicines in line with national guidance. Almost all clinical practice is now mediated through an EHR system; further quantitative research into the effect of EHR system design on clinical practice is long overdue.

Structural Basis for Diltiazem Block of a Voltage-Gated Ca2+ Channel.

Diltiazem is a widely prescribed Ca2+ antagonist drug for cardiac arrhythmia, hypertension, and angina pectoris. Using the ancestral CaV channel construct CaVAb as a molecular model for X-ray crystallographic analysis, we show here that diltiazem targets the central cavity of the voltage-gated Ca2+ channel underneath its selectivity filter and physically blocks ion conduction. The diltiazem-binding site overlaps with the receptor site for phenylalkylamine Ca2+ antagonist drugs such as verapamil. The dihydropyridine Ca2+ channel blocker amlodipine binds at a distinct site and allosterically modulates the binding sites for diltiazem and Ca2+ Our studies resolve two distinct binding poses for diltiazem in the absence and presence of amlodipine. The binding pose in the presence of amlodipine may mimic a high-affinity binding configuration induced by voltage-dependent inactivation, which is favored by dihydropyridine binding. In this binding pose, the tertiary amino group of diltiazem projects upward into the inner end of the ion selectivity filter, interacts with ion coordination Site 3 formed by the backbone carbonyls of T175, and alters binding of Ca2+ to ion coordination Sites 1 and 2. Altogether, our results define the receptor site for diltiazem and elucidate the mechanisms for pore block and allosteric modulation by other Ca2+ channel-blocking drugs at the atomic level. SIGNIFICANCE STATEMENT: Calcium antagonist drugs that block voltage-gated calcium channels in heart and vascular smooth muscle are widely used in the treatment of cardiovascular diseases. Our results reveal the chemical details of diltiazem binding in a blocking position in the pore of a model calcium channel and show that binding of another calcium antagonist drug alters binding of diltiazem and calcium. This structural information defines the mechanism of drug action at the atomic level and provides a molecular template for future drug discovery.

Clinical Safety Outcomes in Patients With Nonvalvular Atrial Fibrillation on Rivaroxaban and Diltiazem.

BACKGROUND: It is unknown whether diltiazem, a moderate cytochrome P450 enzyme (CYP3A4) and P-glycoprotein (P-gp) inhibitor, increases the incidence of bleeding events in combination with rivaroxaban, a CYP3A4 and P-gp substrate. OBJECTIVE: To assess major and clinically relevant nonmajor (CRNM) bleeding outcomes in patients with nonvalvular atrial fibrillation (NVAF) on rivaroxaban with concomitant diltiazem in a real-world setting. METHODS: This retrospective case-cohort study included adult patients with NVAF prescribed both rivaroxaban and diltiazem for at least 30 days. Patients were matched 1:1 by age and baseline creatinine clearance (CrCl) to control patients taking rivaroxaban alone. The primary outcome was the composite of major and CRNM bleeding. Additional outcomes included bleeding events resulting in discontinuation of rivaroxaban, time to first bleeding event, and type of first bleed. RESULTS: A total of 143 cases and 143 controls were included. The mean age was 69 years and median baseline CrCl was 87 mL/min. Median follow-up time was 12.4 months for cases and 16.5 months for controls. There was no significant difference in proportion of patients experiencing a major and/or CRNM bleeding event between cases and controls: 23.1% versus 28.0%, respectively; 9 cases and 8 controls permanently discontinued rivaroxaban because of bleeding. Gastrointestinal/rectal bleeding and hematuria were the most frequently reported bleeding events in both groups. Conclusion and Relevance: This is the first study to assess major and CRNM bleeding outcomes in patients with NVAF on rivaroxaban and diltiazem. Diltiazem use was not associated with an increased rate of bleeding events.

Preventative effect of diclofenac sodium and/or diltiazem in rats with epidural fibrosis.

OBJECTIVE: Spinal epidural fibrosis is commonly seen after laminectomy. There is not yet proven any agent preventing fibrosis in clinical usage. We used diclofenac sodium and diltiazem, which are fibrosis inhibitors. METHODS AND MATERIALS: 40 rats were divided into four groups of equal numbers: control, diclofenac sodium, diltiazem, and diclofenac sodium + diltiazem. Laminectomies were performed at L5 and L6. After a 4 week period, the rats were decapitated and the vertebral column blocks were removed for histopathologic examination. Fibrosis percentage, spread of fibrous regions, and fibroblast numbers were evaluated in each group and compared between the groups. RESULTS: The distribution of epidural fibrosis density, percentage of fibrosis, and distribution of fibroblasts in the diclofenac sodium + diltiazem group were significantly lower than in the other groups. The fibroblast numbers of the diltiazem, and diclofenac sodium + diltiazem groups were significantly lower than in the other groups. CONCLUSION: Diclofenac sodium + diltiazem used together provided better outcomes because each of them prevented fibrosis via different ways, probably through synergistic action (Tab. 5, Fig. 3, Ref. 43).